Mesenchymal stem cells are a multipotent stem cells discovered in bone marrow. Since mesenchymal stem cells have an ability to differentiate into a variety of cell types such as bone cells, cardiac muscle cells, fat cells or the like, it is expected that they can be applied in regenerative medicine in such a manner that they are administered (by injection) to a living body and allowed to transfer to the affected part and differentiate into a tissue needed there (cf. patent document 1 and patent document 2). Also, it has been shown that mesenchymal stem cells can modulate T cell-mediated immune reactions when administered to a living body, and that the cells can be used as a medicine which inhibits rejection after transplantation (cf. patent document 3).
Focusing on the multipotency of and the effect to inhibit rejection by mesenchymal stem cells, clinical studies using human mesenchymal stem cells are going on. For example, led by the U.S. groups, clinical studies are being performed on the therapeutic effect of human mesenchymal stem cells which were collected from a person genetically different from the patients and cultured, on the graft versus host disease (GVHD) occurring after bone marrow transplantation, and their efficacy is being proved (cf. non-patent document 1 and non-patent document 2). Also, a clinical application of human mesenchymal stem cells for myocardium reproduction after occurrence of myocardial infarction is attempted, too (cf. non-patent document 3).
It is known that human mesenchymal stem cells occur in bone marrow fluid. However, their number is very limited. While it is also known that human mesenchymal stem cells are available from various tissues such as adipose tissue (cf. patent document 4), dental pulp tissue (cf. patent document 5), placenta tissue or umbilical cord tissue as well as from bone marrow fluid (cf. patent document 6), the available number of them are very small in any case.
As noted above, human mesenchymal stem cells occurring in the living body are very few. However, a large number of human mesenchymal stem cells can be prepared from a very limited number of human mesenchymal stem cells obtained from bone marrow, through their culture and multiplication using an artificial culture medium (cf. patent document 7). Human mesenchymal stem cells prepared in large numbers in this way can, like other cultured cells, be preserved in a frozen state, and it is known that a frozen culture medium comprising 90% of fetal bovine serum and 10% of dimethyl sulfoxide (DMSO) can be used for cryopreservation (cf. patent document 8). In order for human mesenchymal stem cells to be widely supplied as pharmaceutical products to medical institutions for the treatment of GVHD, myocardium reproduction or the like, it is expected that human mesenchymal stem cells must be transported to the medical institutions in the form of packaged frozen cells to save the cells from possible alteration until the use thereof.
Thus, in order to supply human mesenchymal stem cells in the market as parenteral pharmaceutical products to be administered directly to humans, it is necessary for the products to pass through processes of washing the cells to remove the culture medium used for proliferation thereof and thereafter freezing the cells in a solution which is suitable for the dosage to humans. Generally, each of washing and freezing of the cells, and thawing them before use thereafter gives a major stress to the cells, and some of them are thereby ready to be injured to death. When human mesenchymal stem cells are supplied as pharmaceutical products in a frozen state, it is undesirable, from the viewpoints of maintaining viable cell counts necessary for achieving their pharmaceutical effect, and of their quality control as pharmaceutical products as well, to allow to cause a large number of cell deaths during the processes of preparing frozen cells as pharmaceutical products from cultured cells and of thawing them before use thereafter.
Also, when human mesenchymal stem cells are administered to a patient by intravenous injection, it can be assumed that the cells, after thawed, are added to the infusion fluid in a bag. Therefore, it is desirable that the cells are maintained stable even in the state in which they are, after thawed, added to other infusion fluid and thereby diluted.    [patent document 1] Japanese Patent Application Publication No. H10-512756    [patent document 2] Japanese Patent Application Publication No. 2002-511094    [patent document 3] U.S. Pat. No. 6,328,960    [patent document 4] Japanese Patent Application Publication No. 2004-129549    [patent document 5] Japanese Patent Application Publication No. 2004-201612    [patent document 6] Japanese Patent Application Publication No. 2004-210713    [patent document 7] U.S. Pat. No. 5,486,359    [patent document 8] Japanese Patent Application Publication No. H07-500001    [non-patent document 1] Transplantation. 2006; 81(10): 1390-7    [non-patent document 2] Br J Haematol. 2007; 137(2): 87-98    [non-patent document 3] Arch Iran Med. 2007; 10(4): 467-73